The purpose of this project is the study of Aleutian disease of mink (AD), a persistent infection by the Aleutian disease virus (ADV), a nondefective parvovirus. In the past year, we have utilized reagents and probes to begin detailed study of AD, both in vivo andd in vitro. Using ADV-G infected Crandell feline kidney (CRFK) cells as an in vitro model, we have quantitated infectious virus, viral antigens (both by fluorescence and radioimmunoassay) and viral DNA (by dot-blot hybridization). The data generated by these experiments have allowed us to conclude that an infected CRFK cell expressing ADV antigens contains about 100,000 genome copies of viral DNA. Relating this information to data on viral DNA content of tissues of infected mink, we have determined that at the height of viral replication (10 days after infection with Utah I ADV), approximately about 0.02-0.1% of the cells in mesenteric lymph node, spleen and liver contain virus. In addition, analysis of Southern blot hybridization has localized ADV DNA with characteristics of replicative forms in these tissues, implying they are true sites of replication. Additional studies suggest that the virus is located in cells of the lymphocyte series and not (exclusively) present in phagocytic cells. Current studies are focussing on cells isolated from mesenteric lymph node, and we are presently analyzing lymphocyte populations highly enriched for "T" or "B" lymphocytes. Additional work has suggested that the small MW proteins found in ADV isolated from mink organs are the result of in vivo proteolysis. It is likely that a trypsin-like enzyme cleaves the about 85K and about 75K structural proteins into highly antigenic proteins with MW's between 20-30K, similar to those seen with virus purified from mink organs. These findings will be pursued during the next year by comparative peptide mapping of in vivo and in vitro preparations of ADV.